Although environmental and chemical carcinogens are considered to be important in the causation of human cancer, and polycyclic hydrocarbons, nitrosoamides, aromatic amine derivatives and nitroquinolines induce breast cancer in experimental animals, they have not been proven to be carcinogenic for human breast epithelial cells (HBEC). Assessment of the role of chemical carcinogens in human breast cancer requires the understanding of the mechanisms by which normal cells become malignant and the availability of a reproducible, quantitative system for transforming HBEC by exposure to carcinogens. In vitro treatment of HBEC with the chemical carcinogens DMBA and NMU induces phenotypic changes, such as anchorage independent growth and multinucleation, which are modulated by the degree of gland development and pregnancy history. Despite these advances, neither our group nor other investigators have, as yet, successfully developed a reproducible system for the complete transformation of HBEC into malignant cells with chemical carcinogens. This could be due to inadequacy of the models utilized for HBEC transformation, to the fact that chemical carcinogentreated cells expressing in vitro phenotypic changes of cell transformation require an adequate heterologous system for expression of the malignant phenotype, or that genotoxic chemical carcinogens require an inherently susceptible target cell. In order to determine which factor(s) is/are responsible for the modulation of the susceptibility of HBEC to malignant transformation, we propose: a) to use culture conditions that maintain cells for long periods of time without terminal differentiation or senescence, and that permit identification of the transformed phenotype; b) to identify the phenotypes that indicate early and late events of cell transformation, characterizing each of the multisteps involved in carcinogenesis; c) to determine whether cells exhibiting phenotypic markers of cell transformation in vitro express in a heterologous host definitive malignant phenotypes such as tumorigenicity, and d) to uncover which common and constant features affecting the susceptibility of the mammary gland to carcinogenesis are exhibited by the heterogeneous human population, and whether those features are a function of age or reproductive history. The answer to these questions will have immediate application to the development of control measures for protecting the human breast from neoplastic transformation.